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Purification of Protein-complexes from the Cyanobacterium Synechocystis sp. PCC 6803 Using FLAG-affinity Chromatography

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Analyzing the Quenchable Iron Pool in Murine Macrophages by Flow Cytometry

Authors: Michael Riedelberger and Karl Kuchler, date: 03/20/2020, view: 408, Q&A: 0

[Abstract] Tissue-resident macrophages are pivotal for a tightly-regulated iron metabolism at a cellular and systemic level, since subtle iron alterations increase the susceptibility for microbial infections or drive multiple diseases. However, research on cellular iron homeostasis in macrophages remains challenging due to the limited amount of available ...

Propagation and Purification of Chlamydia trachomatis Serovar L2 Transformants and Mutants

Authors: Robert Faris and Mary M. Weber, date: 12/20/2019, view: 741, Q&A: 0

[Abstract] Chlamydia trachomatis (C.t.) is an obligate intracellular pathogen that cannot be cultured axenically and must be propagated within eukaryotic host cells. There are at least 15 distinct chlamydial serovariants that belong to 2 major biovars commonly referred to as trachoma and lymphogranuloma venereum (LGV). The invasive ...

Lysosome Targeting RedGreen-assay: Selective Autophagy Sensing Assay for Mammalian Cells

Authors: Victoria Riccio, Miluska Vissa, Angus McQuibban and Peter Kijun Kim, date: 12/20/2019, view: 1175, Q&A: 0

[Abstract] The process of autophagy is an essential cellular mechanism, required to maintain general cell health through the removal of dysfunctional organelles, such as the ER, peroxisomes and mitochondria, as well as protein aggregates, and bacteria. Autophagy is an extremely dynamic process, and tools are constantly being developed to study the various ...

Bacterial Synchronized Transfer Assays in Bone Marrow Derived Macrophages

Authors: Sedelia R Dominguez, Tom Kawula and Shaun P Steele, date: 11/20/2019, view: 1026, Q&A: 0

[Abstract] Merocytophagy (“mero”, Greek for partial; “cytophagy” for cell eating) is a process by which cells acquire microbes and cytosolic material through phagocytosis of a small portion of neighboring cells upon cell-cell contact. Cell-cell contact dependent transfer events can be assessed through co-incubation of differently labeled cells. With these ...

Quantitation of TLR4 Internalization in Response to LPS in Thioglycollate Elicited Peritoneal Mouse Macrophages by Flow Cytometry

Authors: Katharina Richard, Stefanie N. Vogel and Darren J. Perkins, date: 09/20/2019, view: 1042, Q&A: 0

[Abstract] Internalization of cell surface Toll-like Receptor 4 (TLR4) is a convenient read-out to measure LPS dependent activation of the TRIF adaptor pathway. We here provide a protocol to quantify the LPS dependent internalization of TLR4 using thioglycollate-elicited peritoneal macrophages by flow cytometry.

Intracellular Invasion and Killing Assay to Investigate the Effects of Binge Alcohol Toxicity in Murine Alveolar Macrophages

Authors: Victor Jimenez Jr and Fernando P Monroy, date: 01/20/2019, view: 2106, Q&A: 0

[Abstract] Alcohol consumption has diverse and well-documented effects on the human immune system and its ability to defend against infective agents. While pulmonary related infections can occur in healthy humans, binge alcohol use is recognized as a major health risk factor (Nelson et al., 1991). Although binge alcohol consumption has been ...

Method for Studying the Effect of Gene Silencing on Bacterial Infection-induced ERK1/2 Signaling in Bone-marrow Derived Macrophages

Authors: Gaurav Kumar, Subhash B. Arya and Amit Tuli, date: 12/20/2018, view: 2466, Q&A: 0

[Abstract] Macrophages are highly phagocytic cells that utilize various pathogen recognition receptors (PRRs) to recognize pathogen-associated molecular patterns (PAMPs). These PAMPs can be present within the microbe, such as bacterial CpG DNA, and are recognized by Toll-like receptor 9 (TLR9), a PRR present on the endosomal membrane of macrophages. PAMPs ...

Phagocytosis Assay for α-Synuclein Fibril Uptake by Mouse Primary Microglia

Authors: Cuilian Du, Feifan Zhang and Claire Xi Zhang, date: 09/05/2018, view: 2593, Q&A: 0

[Abstract] Microglia are professional phagocytes in the brain and deficiency in their phagocytic activity plays an important role in Parkinson’s disease. This protocol mainly describes the phagocytosis assay for uptake of α-synuclein preformed fibrils, a pathologic form of α-synuclein, by primary microglia.

Measurement of TLR4 and CD14 Receptor Endocytosis Using Flow Cytometry

Authors: Michael S. Schappe and Bimal N. Desai, date: 07/20/2018, view: 4765, Q&A: 0

[Abstract] After recognizing extracellular bacterial lipopolysaccharide (LPS), the toll-like receptor 4 (TLR4)-CD14 signaling complex initiates two distinct signaling pathways–one from the plasma membrane and the other from the signaling endosomes (Kagan et al., 2008). Understanding the early stages of TLR4 signal transduction therefore requires a ...

3D Co-culture System of Tumor-associated Macrophages and Ovarian Cancer Cells

Authors: Lingli Long, Mingzhu Yin and Wang Min, date: 04/20/2018, view: 4883, Q&A: 0

[Abstract] Ovarian cancer is fairly unique in that ovarian carcinoma cells can detach and spread directly through peritoneal cavity. It has been unclear, however, how detached cancer cells survive in the peritoneum and form spheroid structure. We have recently reported that there is a strong correlation between Tumor-associated macrophages (TAMs)-associated ...

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